Short- and long-term redox regulation of photosynthetic light energy distribution and photosystem stoichiometry by acetate metabolism in the green alga, Chlamydobotrys stellata.

The effect of acetate metabolism on the light energy distribution between the two photosystems, on the PS II/PS I stoichiometry and on the expression of psbA and psbB and psaA genes was investigated in the green alga, Chlamydobotrys stellata during autotrophic (CO(2)), mixotrophic (CO(2) plus acetate) and photoheterotrophic (only acetate) cultivation. It was observed that acetate assimilation in the glyoxylate cycle resulted in a large drop in the ATP content and a concomitant increase in the NADPH content of the cells. The combined effect of high NADPH concentration and linear electron transport brought about an over-reduction of the inter-photosystem electron transport components. The reduced state of the inter-photosystem components initiated a state 1/state 2 transition of LHC II and a decrease in the PS II/PS I ratio. The PS II/ PS I ratio was reduced because the synthesis of PS II reaction centers was repressed and that of the PS I reaction centers was slightly enhanced by acetate cultivation. The amount of PsbA and PsbB proteins of PS II and the abundance of psbA mRNA decreased. The abundance of PS I PsaA protein and psaAmRNA were only slightly increased. All of the acetate-induced effects were reversible when the cells were transferred back to an acetate-free medium. Our observations demonstrate that the expression of the PS II psbA and psbB and PS I psaA genes is regulated by the redox state of the inter-photosystem components at the transcriptional level. Experiments carried out in the presence of DBMIB which facilitates the reduction of plastoquinone pool indicate that the expression of genes encoding the components of PS II and PS I are controlled by the redox state of a component (cytochrome b/f complex) located behind the plastoquinone pool.

Influence of photoheterotrophy on the expression of chlorophyll a/b-binding proteins in the green alga Pyrobotrys stellata.

Two genes (lhca5 and lhcb1) from the unicellular, green alga Pyrobotrys (formerly Chlamydobotrys) stellata were isolated, coding for Chlorophyll a/b-binding proteins that putatively represent constituents of the light-harvesting complexes connected with Photosystem I and Photosystem II, respectively. Expression of both genes on the mRNA-level is markedly inhibited by CO2-depletion. The lhca5 transcript-level was reduced to about 25%, and the lhcb1-expression was completely blocked 9 h after removal of CO2 from the growth medium. Simultaneous addition of acetate, which can substitute for CO2 as a carbon source during photoheterotrophic growth of P. stellata, did not compensate for the diminishing effect of CO2-depletion on lhcb1. However, the amount of lhca5 transcript was comparable to that during photoautotrophic growth. These results are interpreted in terms of the specific metabolic demands of photoheterotrophic growth in P. stellata. Cyclic electron-transfer along Photosystem I must be sustained for ATP-production. Linear electron transport fed by Photosystem II and concomitant production of NADPH for CO2-reduction is no longer required.The sequences reported in this article have been deposited at the EMBL data library under the accession numbers X69434 (CSCAB1) and X71965 (CSCAB2MR).

Involvement of the donor tyrosine-D1 (Y d) in Photosystem II electron transport in the green alga, Chlamydobotrys stellata.

The light-induced oxidation of the accessory donor tyrosine-D (YD) has been studied by measurements of the EPR Signal IIslow at room temperature in the autotrophically and photoheterotrophically cultivated alga Chlamydobotrys stellata. After illumination and dark adaptation, YD Signal IIslow was observed only in autotrophic algae, i.e. under conditions of a linear photosynthetic electron transfer from water to NADP(+). The addition of artificial electron acceptors phenyl-p-benzoquinone (PPQ) or dichloro-p-benzoquinone (DCQ) to the autotrophic cells caused an almost negligible increase of this signal. When photosynthetic electron flow and oxygen evolution were diminished by removal of the carbon source CO2 and addition of acetate (photoheterotrophy), a pronounced YD Signal IIslow was seen only in presence of DCQ or PPQ. Several possibilities are discussed to explain the absence of YD Signal IIslow in photoheterotrophic Chl. stellata such as the existence of a cyclic PS II electron flow very effectively reducing P680 and thereby preventing the possibility of YD oxidation. Artificial electron acceptors withdraw electrons from this cycle thus keeping the primary quinone acceptor, QA, oxidized and thereby diminishing the reduction of P680 (+) by cyclic PSII. This leads to the appearance of the YD Signal IIslow also in the photoheterotrophically grown algae.

Treatment of sepsis and septic shock: a review.

Septic shock is one of the leading causes of death in intensive care units, and its incidence is increasing. Mortality rates as high as 95% are reported, with rates of 60% or more even when diagnosed and treated promptly. This review examines the definition of septic shock, its pathogenesis, and supportive therapy, with particular attention to intervention during the septic shock cascade.

Hydrogen Photoevolution Indicates an Increase in the Antenna Size of Photosystem I in Chlamydobotrys stellata during Transition from Autotrophic to Photoheterotrophic Nutrition.

The changes in the light-harvesting antenna size of photosystem I were investigated in the green alga Chlamydobotrys stellata during transition from autotrophic to photoheterotrophic nutrition by measuring the light-saturation behavior of hydrogen evolution following single turnover flashes. It was found that during autotrophic-to-photoheterotrophic transition the antenna size of photosystem I increased from 180 to 250 chlorophyll. The chlorophyll (a + b)/P700 ratio decreased from 800 to 550. The electron transport of photosystem I measured from reduced 2,6-dichloro-phenolindophenol to methylviologen was accelerated 1.4 times. In the 77K fluorescence spectra, the photosystem II fluorescence yield was considerably lowered relative to the photosystem I fluorescence yield. It is suggested that the increased light-harvesting capacity and redistribution of absorbed excitation energy in favor of photosystem I is a response of photoheterotrophic algae to meet the ATP demand for acetate metabolism by efficient photosystem I cyclic electron transport when the noncyclic photophosphorylation is inhibited by CO(2) deficiency.

Thermoluminescence study of the in vivo effects of bicarbonate depletion and acetate/formate presence in the two algae Chlamydobotrys stellata and Chlamydomonas reinhardtii.

We investigated the influence of CO2/HCO3 (-)-depletion and of the presence of acetate and formate on the in vivo photosynthetic electron transport in the two green algae Chlamydobotrys stellata and Chlamydomonas reinhardtii by means of thermoluminescence technique and mathematical glow curve analysis. The main effects of the removal of CO2 from the algal cultures was: (1) A shift of the glow curve peak position to lower temperatures resulting from a decrease of the B band and an increase of the Q band. (2) Treatment of CO2-deficient Chl. stellata with DCMU yielded two thermoluminescence bands in the Q band region peaking at around +12°C and +5°C; in case of Chl. reinhardtii DCMU treatment induced only one band with an emission maximum at +5°C. The presence of acetate or formate in CO2-depleted algal cultures lowered the intensities of all of the individual TL bands but that of a HT band (TL+37). The effects of CO2-depletion and of the presence of anions were fully reversible.

Flash oxygen yield patterns of autotrophically and photoheterotrophically grown Chlamydobotrys stellata in the presence and absence of lipophilic acceptors.

The obligate phototrophic green alga Chlamydobotrys stellata does not evolve oxygen when grown in CO2-free atmosphere on acetate. With the application of the lipophilic acceptor 2,6-dichloro-p-benzoquinone it was investigated whether this phenomenon is caused by the inactivation of the water-splitting system or by an inhibition of the electron transport chain. It was found that in the presence of DCQ, the photoheterotrophic alga exhibited a normal period-4 flash oxygen pattern, but the steady state yield was only 25% of that measured in the autotrophic cells. After DCQ addition, the initial distribution of S-states and the values of the transition probabilities proved to be the same in the autotrophic and photoheterotrophic algae. These results indicate that photoheterotrophic growth conditions inhibit the electron transport of Chl. stellata behind the acceptor site of DCQ, but the water-splitting system remains active with a reduced oxygen evolving capacity.

Infection of algae-free Paramecium bursaria with symbiotic Chlorella sp. Isolated from green paramecia: I. Effect of the incubation period.

The significance of the length of incubation (30 sec to 48 h) of algae-free Paramecium bursaria with symbiotic Chlorella sp. for the success of infection, i.e. the reestablishment of the endosymbiotic algae has been investigated. When algae are brought together with paramecia, they are rapidly taken up by the ciliates. During a 30 sec incubation one ciliate engulfs about 50 chlorellae. A prolongation of the incubation period increases the number of ingested algae. However, the success of infection, determined one and five day(s) after the end of the incubation, is independent from the length of the incubation period and, consequently, does not depend on the number of ingested algae, either: In all experiments about 50% of the Paramecium population becomes infected and one to three algae are primarily enclosed in individual perialgal vacuoles within a ciliate cell. Thus, the endosymbiont population of a Paramecium cell originates on an average from two algae. Since successful infection is restricted only to a part of the Paramecium population and since the number of primarily established endosymbionts does not depend on the number of ingested algae, the success of infection and the formation of perialgal vacuoles seem to be not limited by properties of the algae but by features of the host, the possible nature of which is discussed.

Comparative thermoluminescence study of autotrophically and photoheterotrophically cultivated Chlamydobotrys stellata.

Thermoluminescence (TL) from autotrophically and photoheterotrophically cultivated Chlamydobotrys stellata was measured. Strong TL was emitted at 30°C after acetatenutrition of the alga. DCMU enhanced this band, as also did ferricyanide. It also appeared after poisoning of the alga with NH2OH or ANT-2p. These observations suggest that an alternative donor to photosystem II and not the water-splitting system is responsible for the TL + 30 band.

Bicarbonate in vivo Requirement of Photosystem II in the Green Alga Chlamydobotrys stellata.

Flash induced 685 nm fluorescence emission of preilluminated and dark kept Chlamydobotrys stellata has been measured under conditions of CO(2)-deprivation. The difference in fluorescence intensity between dark kept and preilluminated cells is taken as a measure for the reduced state of the primary stable electron acceptor of photosystem II, Q, at the given intensity of preillumination. CO(2) removal from growing cultures of this alga for 15 min diminishes photosynthetic electron transport at the oxidizing side of this photosystem. Prolonged CO(2)-absence influences also its reducing side. Measurements of flash induced oxygen yields support the conclusion that both sides of photosystem II are affected in the absence of bicarbonate.

Comparative freeze-fracture study of perialgal and digestive vacuoles in Paramecium bursaria.

In the endosymbiotic unit of Paramecium bursaria (Ciliata) and Chlorella sp. (Chlorophyceae) algae are enclosed individually in perialgal vacuoles, which do not show acid phosphatase activity and thus differ from digestive vacuoles. Both types of vacuoles have been studied by freeze-fracture. Perialgal vacuoles are nearly spherical; their membrane always fits tightly to the algal surface. The vacuole size and shape do not vary much. During division of the algal cell into four autospores the vacuole diameter only doubles. After autospore formation the vacuole invaginates around the algal daughter cells and divides. Newly formed perialgal vacuoles remain in intimate contact and exhibit characteristic attachment zones before final separation. The two fracture faces of perialgal vacuole membranes are homogeneously covered with intramembranous particles (IMPs) but rarely show signs of vesicles pinching off or fusing with the membrane, except during vacuole division. The P-faces bear more IMPs (3164 +/- 625 IMP/micron 2) than the E-faces (654 +/- 208 IMP/micron 2). The range of IMP density on both faces is enormous, suggesting that the membrane is not static. Membrane changes are supposed to occur simultaneously with the enlargement of the vacuole and to be caused by fusion with cytoplasmic vesicles, as the fractured necks on vacuole membranes may indicate. Digestive vacuoles in P. bursaria show significant variations in size, shape, membrane topography and IMP density, as well as signs of endocytic activity. Different vacuole populations are present in P. bursaria according to different feeding conditions: ciliates fed for a long time have small vacuoles with few IMPs (322 +/- 198 IMP/micron 2 on the E-faces, 1438 +/- 458 IMP/micron 2 on the P-faces), which are probably condensed digestive vacuoles, whereas organisms fed for a short time have larger vacuoles with highly particulate faces (680 +/- 282 IMP/micron 2 on the E-faces, 2701 +/- 503 IMP/micron 2 on the P-faces) and thus are supposed to be older vacuoles. The digestive vacuole membrane changes continuously. Compared to digestive vacuoles perialgal vacuoles are characterized by small size combined with high IMP density on the two fracture faces. Their IMP densities resemble those of old digestive vacuole membranes. However, it would be premature to conclude that membranes of perialgal and old digestive vacuoles are identical. Membranes of old digestive vacuoles are mainly derived from lysosomal material, which presumably does not contribute to the formation of perialgal vacuole membranes as is indicated by the small vacuole diameter; fusion with lysosomes would considerably enlarge it.(ABSTRACT TRUNCATED AT 400 WORDS)

Induction of the De Novo Formation of the Photosystem I-Related Light-Harvesting Chlorophyll Protein Complex LHCPa by Photoheterotrophic Nutrition.

The green alga Chlamydobotrys stellata contains in addition to the normal light-harvesting chlorophyll protein complex LHCPb a special LHCPa which is free of chlorophyll b and connected only to photosystem I (Brandt, Zufall, Wiessner 1983 Plant Physiol 71: 128-131). The kinetics of these two LHCP forms were analyzed during the transition in nutrition of the alga from autotrophy to photoheterotrophy, e.g. the replacement of CO(2) by acetate as carbon source. As shown by incorporation experiments with [(14)C]acetate, this change in nutrition leads to an increased synthesis of LHCPa, whereas the synthesis of the photosystem II-related LHCPb decreases. The increase of the LHCPa synthesis starts immediately after the onset of photoheterotrophic nutrition together with the synthesis of the chlorophyll protein complex CPI. There is no interchange of complex components between LHCPa and LHCPb during the depletion of the latter. The formation of LHCPa is discussed with respect to the regulation of gene expression.

Relation between the Light-Harvesting Chlorophyll a-Protein Complex LHCPa and Photosystem I in the Alga Chlamydobotrys stellata.

The light-harvesting chlorophyll protein system of the alga Chlamydobotrys stellata consists of an as yet uncharacterized algal chlorophyll a-protein, called LHCPa, and a common photosystem II-related chlorophyll a/b-protein, called LHCPb (Brandt, Kaiser-Jarry, Wiessner 1982 Biochim Biophys Acta 679: 404-409). For further characterization, this LHCPa was isolated from the organism by polyacrylamide isoelectrofocusing and reelectrophoresis. It contains only chlorophyll a and has only one apoprotein (32,000 daltons). When separated from autotrophically grown cells, its absorption peak is at 674 nm and its isoelectric point at 5.3. Photoheterotrophic cultivation of the algae shifts the absorption maximum of LHCPa to 679 nm and its isoelectric point to 4.8. This LHCPa is a component of photosystem I particles. In relation to the total chlorophyll a content, the amount of LHCPa is low in autotrophic algae, but increases under photoheterotrophic growth conditions, where the organisms do not have the ability to assimilate CO(2) photosynthetically.

Participation of algal surface structures in the cell recognition process during infection of aposymbiotic Paramecium bursaria with symbiotic chlorellae.

The endosymbiotic unit green Paramecium shows a strong specificity of its partners. The aposymbiotic Paramecium bursaria forms a stable symbiotic unit only with a special strain of Chlorella sp. Algae suitable for symbiosis formation are enclosed in individual perialgal vacuoles whereas unsuitable algae are sequestered into food vacuoles. It is probable that algae are recognized by the ciliate because of specific surface structures rather than by their physiological properties. Experiments with synchronized algae demonstrate that autospores are taken up into perialgal vacuoles to a higher degree than mother cells, which have a different surface structure as shown by immunological techniques. Symbiotic algae treated with cellulase and pectinase or having been coated with specific antibodies or with lectins (concanavalin A or Ricinus communis agglutinin) are usually not recognized as suitable and are mostly sequestered into food vacuoles although they show the same physiological properties as untreated algae. These results indicate the participation of carbohydrate structures at the recognition sites of symbiotic chlorellae in Paramecium bursaria which interact during infection with special receptor molecules in the membrane of the ingestion vacuole of the ciliate.

On the developmental dependence between Cyanophora paradoxa and Cyanocyta korschikoffiana in symbiosis : Host-dependent development of endocyanelles.

The prokaryote Cyanocyta korschikoffiana was isolated from the eukaryote Cyanophora paradoxa. The synthesis of several thylakoid proteins in these cyanelles is influenced by light and darkness and is sensitive to cycloheximide, the inhibitor of the eukaryotic host's translation. The possibility of a direct coordination between the translations of the host and of the cyanelles is discussed.

Photobehaviour of Paramecium bursaria infected with different symbiotic and aposymbiotic species of Chlorella.

The endosymbiotic unit of Paramecium bursaria and Chlorella spec. shows two types of photobehaviour: 1) A step-up photophobic response which possibly depends on photosensitive agents in the ciliate cell itself - as is also shown by alga-free Paramecium bursaria - and can be drastically enhanced by photosynthetic activity of symbiotic algae; and 2) a step-down photophobic response. The step-down response leads to photoaccumulation of green paramecia. Both types of photobehaviour in Paramecium bursaria do not depend on any special kind of algal partners: The infection of alga-free Paramecium bursaria with different Chlorella species results in new ciliatealgae-associations. They are formed not only by combination of the original symbiotic algae with their host, but also by infection with other symbiotic or free-living (aposymbiotic) chlorellae, respecitively. Systems with other than the original algae are not permanently stable - algae are lost under stress conditions - but show the same types of photobehaviour. Photoaccumulation in general requires algal photosynthesis and occurs only with ciliates containing more than fifty algae/cell. It is not mediated by a chemotactic response to oxygen in the medium, since it occurs at light fluence rates not sufficient for a release of oxygen by the symbiotic system, e.g., below its photosynthetic compensation point. Photoresponses can be inhibited by 3-(3',4'-dichlorophenyl)-1,1-dimethylurea (DCMU). Sensory transduction does not depend on any special symbiotic features of the algae, e.g., sugar excretion. The participation of oxygen in the Paramecium cell, of its cytoplasmic pH and of ions released or taken up by endosymbiotic algae in sensory transduction is discussed.

Evidence of de novo synthesis of maltose excreted by the endosymbiotic Chlorella from Paramecium bursaria.

The endosymbiotic Chlorella sp. from Paramecium bursaria excretes maltose both in the light and in the dark. Experiments on photosynthetic (14)CO2 fixation and (14)CO2 pulse-chase experiments show that maltose is synthesized in the light directly from compounds of the Calvin cycle, whereas in the dark it results from starch degradation.

The role of endosymbiotic algae in photoaccumulation of green Paramecium bursaria.

The endosymbiotic unit of Paramecium bursaria with Chlorella sp. photoaccumulates in white, blue-green, and red light (λ<700 nm), whereas alga-free Paramecia never do. The intensity of photoaccumulation depends on both the light fluence rate and the size of the symbiotic algal population. Photoaccumulation can be stopped completely with 3-(3',4'-dichlorophenyl)-1,1-dimethylurea (DCMU), an inhibitor of photosynthetic electron transport. Hence the photosynthetic pigments of the algae act as receptors of the light stimulus for photomovement and a close connection must exist between photosynthesis of the algae and ciliary beating of the Paramecium.